DESCRIPTION (Abstract reproduced verbatim): The goal of this project is to understand the role of nitric oxide (NO) as a regulator of cell proliferation and differentiation. Our studies have shown that NO acts as an anti proliferative agent during organism development, regulating the balance between cell proliferation and differentiation in the differentiating tissue, and, ultimately, controlling the shape and size of tissues and organs in the developing organism. Thus, NO emerges as an essential negative regulator of cell proliferation and a mediator of tissue differentiation. In this project, we will study the mechanisms of the anti proliferative action of NO during cell division and differentiation and identify the molecular targets of NO within the cell cycle machinery. In our first Specific Aim we will start by defining the phases of the cell cycle where the action of NO is most pronounced. Next we will dissect the crucial pathways within each of these phases and identify the stages that are affected by NO. After that, we will probe the individual potential targets of NO. In our second Specific Aim we will investigate whether cell cycle-related signaling by NO synthases is different from signaling induced by exogenous NO donors. Furthermore, we will study the signaling induced by each of the three NOS isoforms under similar conditions in order to identify the isoform-specific targets of NOS. In our third Specific Aim we will test the hypothesis that most of the antiproliferative activity of NGF during neuronal differentiation of PC12 cells is mediated by NOS. For this, we will determine the genes induced by NGF in PC12 cells and compare them to the genes induced by NO donors and NOS gene, will determine which of the genes induced by NGF are dependent on NOS activity for their activation, and will investigate whether NGF induced activation of p53 is dependent on NO production.